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Radwan and Ahmed Diagnostic Pathology 2012, 7:149 
http://www.diagnosticpathology.org/content/771/149 



DIAGNOSTIC PATHOLOGY 



RESEARCH Open Access 



The diagnostic value of arginase-1 
immunostaining in differentiating hepatocellular 
carcinoma from metastatic carcinoma and 
cholangiocarcinoma as compared to HepPar-1 

Nehal A Radwan" and Naglaa S Ahmed 
Abstract 

Background: The ability to distinguish hepatocellular carcinoma (HCC) from metastatic carcinoma (MC) involving 
the liver and cholangiocarcinoma (CC) by immunohistochemistry has been limited by the lack of a reliable positive 
marker for hepatocellular differentiation. Arginase-1 is a marker for HCC recently described in some literature. 

Aim: To examine the immunohistochemical staining of arginase-1 in cases of HCC, MC involving the liver and CC 
as compared to hepatocyte paraffin antigen -1 (HepPar-1) in an attempt to further define the diagnostic utility of 
arginase-1 in differentiating these tumors. 

Materials and methods: A comparative immunohistochemical study of arginase-1 and HepPar-1 expression was 
performed in 50 HCC cases, 38 cases of MC to the liver from varying sites, 12 cases of CC and 10 specimens of 
normal liver tissues. The predictive capacity of arginase-1 and HepPar-1 staining was determined using sensitivity, 
specificity, positive predictive value, and negative predictive value calculations. 

Results: All normal liver tissues (no=10), non- neoplastic cirrhotic liver tissues adjacent to HCC (no=42) as well as 
those adjacent to MC (no= 9) showed diffuse and strong immunostaining for both arginase-1 and HepPar-1. 
Arginase-1 demonstrated positive immunoreactivity in 42 of 50 (84%) cases of HCC compared with 35 of 50 (70%) 
for HepPar-1. Only one of 38 (2.6%) cases of MC and one of 12 (8.3%) cases of CC showed positive 
immunoreactivity for arginase-1. In contrast, HepPar-1 immunoreactivity was detected in 6 of 38 (15.8%) cases of 
MC and in 2 of 12 (16.7%) cases of CC. Arginase -1 showed a significantly higher sensitivity for HCC diagnosis 
(84%) compared to HepPar -1(70%) (p=0.016). The specificity of arginase-1 for HCC diagnosis was higher (96%) than 
that of HepPar -1 (84%); nevertheless, this was not statistically significant (p=0.109). Howerver, the combination of 
both immunomarkers for the diagnosis of HCC, raised the specificity to 100%. 

Conclusion: Arginase-1 immunostaining has a higher sensitivity and specificity than HepPar-1 for HCC diagnosis. 
Furthermore, the combined use of arginase-1 and HepPar-1 can provide a potentially promising tool to improve 
the accuracy in distinguishing HCC from metastatic carcinoma and cholangiocarcinoma. 

Virtual slides: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/ 
vs/9991 436558072434. 

Keywords: Arginase-1, HepPar-1, Hepatocellular carcinoma, Metastatic carcinoma, Cholangiocarcinoma 



* Correspondence: neharadwan@yahoo.com 

Pathology Department, Faculty of Medicine, Ain Shams University, Cairo, 
Egypt 

O© 201 2 Radwan and Ahmed; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the 
BiolVlGCl C6ntT8l Creative Commons Attribution License (http://creativecommons.Org/licenses/by/2.0), which permits unrestricted use, 
distribution, and reproduction in any medium, provided the original work is properly cited. 



Radwan and Ahmed Diagnostic Pathology 2012, 7:149 
http://www.diagnosticpathology.org/content/771/149 



Page 2 of 12 



Introduction 

Hepatocellular carcinoma (HCC) is the most common 
primary liver cancer. The annual number of new cases 
of HCC worldwide is over one million. Globally, it is the 
fifth most common cancer and the third leading cause 
of cancer related death, preceded only by the lung and 
stomach cancers [1]. The burden of HCC has been 
increasing in Egypt with a doubling in its incidence rate 
in the past 10 years [2]. HCC contributes to 14.8% of all 
cancer mortality in Egypt. It is the second most frequent 
cancer type in Egyptian males after bladder cancer. The 
high incidence of HCC in Egypt is attributed to the 
high prevalence of hepatitis C virus (HCV). HCV is cur- 
rently the most significant public health problem in 
Egypt with an overall prevalence of 17.4% in males and 
12.2% in females [3]. 

The distinction of HCC from cholangiocarcinoma and 
other types of adenocarcinoma metastatic to the liver is a 
relatively frequent, often challenging dilemma for surgical 
pathologists and very crucial, as the treatment goal for 
these tumors are different. Several treatment modalities, 
including surgical resection, radiofrequency ablation, and 
transarterial chemoembolization/radioembolization, are 
available for hepatocellular carcinoma. In contrast, the 
therapeutic approach for metastatic carcinoma of the 
liver is often palliative. Thus, correct classification of 
these tumors is critically important. Although in most 
cases; the correct diagnosis can be reached through a 
synthesis of clinical findings, diagnostic imaging modal- 
ities and routine evaluation of hematoxylin and eosin 
(H&E) stained sections, immunohistochemistry may play 
a very valuable role in clinically atypical and pathologic- 
ally indeterminate cases, especially challenging because 
limited tissue is available with core biopsies, so an appro- 
priate selection of antibodies is imperative [4,5]. 

A limited number of diagnostically useful immuno- 
histochemical markers for identification of hepatocytes 
in routine surgical pathology practice are available in- 
cluding; hepatocyte paraffin antigen-l(HepPar-l), poly- 
clonal carcinoembryonic antigen (CEA), and CD10, with 
alfa-fetoprotein (AFP) and glypican-3 labeling some 
HCCs [6]. However, the utility of each of these markers 
is limited either by suboptimal sensitivity or difficulty in 
interpretation [7]. For example, AFP suffers from a low 
sensitivity of 30% to 50% and its frequent focal staining 
limiting its utility in small biopsy samples [7-10]. Poly- 
clonal CEA and CD 10 can be difficult to interpret be- 
cause canalicular and diffuse cytoplasmic staining can be 
difficult to distinguish. Furthermore, the sensitivities of 
these markers can be low (25% to 50%) in poorly differ- 
entiated HCCs for polyclonal CEA and 50% for CD10) 
[8,10,11]. Over the past decade, HepPar-1, a mitochon- 
drial urea cycle antigen, has been increasingly used as a 
positive marker for hepatic differentiation. [7,9,12-14]. 



However, HepPar-1 also suffers from relatively low sensi- 
tivity in poorly differentiated hepatocellular carcinomas, 
where the distinction between hepatocellular carcin- 
oma and adenocarcinoma is most difficult [9,10,13]. 
In addition, whereas most adenocarcinomas are negative 
for HepPar-1, gastric, esophageal, and pulmonary adeno- 
carcinomas can demonstrate strong cytoplasmic HepPar-1 
staining [7,9,13]. Glypican-3, a heparin sulphate proteogly- 
can expressed at high levels in HCC, has shown high 
specificity with suboptimal sensitivity in the diagnosis of 
HCC when used in isolation as it is well known to be 
immunoreactive in a wide variety of tumors, including 
pulmonary squamous cell carcinoma, [15] germ cell 
tumors, [16] and a subset of gastric adenocarcinomas [17]. 

A recent literature report characterized a new immu- 
nohistochemical marker, arginase-1 as a potential marker 
of hepatocellular differentiation in both surgical path- 
ology and cytopathology. Arginase exists in 2 isoforms, 
namely arginase- 1 and arginase-2, both of which are 
responsible for the hydrolysis of arginine to ornithine 
and urea in the urea cycle. Of the 2 isoforms, arginase-1 
demonstrates high levels of expression within the liver, 
whereas arginase-2 levels are highest in the kidneys and 
pancreas and are very low in the liver [6,18]. Arginase-1 
is expressed in normal human liver with a high degree 
of specificity [19]. Specifically, it has been shown by 
immunohistochemistry to be concentrated in periportal 
hepatocytes [20]. 

The current study aims to examine the immuno- 
histochemical staining of arginase-1 in cases of HCC, 
metastatic carcinoma involving the liver and cholan- 
giocarcinoma as compared to HepPar-1 that is conven- 
tially used. This is in an attempt to further define the 
diagnostic utility of arginase-1 as a reliable positive 
marker in differentiating these tumors. 

Materials and methods 

Tissue collection 

This retrospective study consisted of 50 cases of hepato- 
cellular carcinoma, 38 cases of metastatic carcinoma to 
the liver, 12 cases of cholangiocarcinoma and 10 speci- 
mens of normal liver tissues. All cases were retrieved 
from the archives of the Pathology Department, Ain 
Shams University Hospitals during the period between 
2006 and 2011. The clinical history, pathology reports 
and hematoxylin and eosin (H&E) stained slides for all 
cases were reviewed to confirm the diagnosis. The histo- 
logic grade of HCC was established using the World 
Health Organization criteria [21]. The study was carried 
out with full local ethics approval. 

Immunohistochemical staining procedure 

Four - micron thick sections of the formalin-fixed, 
paraffin-embedded tissue blocks of all the studied cases 



Radwan and Ahmed Diagnostic Pathology 2012, 7:149 
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Page 3 of 12 



were investigated for the presence of a rabbit polyclonal 
antibody against arginase-1 (H-52: sc 20150, Santa Cruz, 
Europe) at a dilution 1:200, and a mouse monoclonal 
antibody against Hep Par-1, (clone OCH1E5, MS-1810- 
R7, ready to use, Lab vision, CA, USA) with a labelled 
streptavidin- biotin-peroxidase complex technique. Briefly, 
Tissue sections were deparaffinized and hydrated in 
xylene and descending grades of alcohol After rinsing 
in PBS, antigen retrieval was performed by treating the 
tissue sections with citrate buffer, pH 6.0 for 10 min 
in a 700-W microwave oven. The endogenous peroxidase 
activity was blocked by incubating the slides in 3% hydro- 
gen peroxide for 5 to 10 min, and then washed in buffer. 
This is followed by incubation with the primary antibody 
(arginase-1 or HepPar-1) for 1 h at room temperature. 
The antibody reaction was detected with the avidin-biotin 
detection kit using diaminobenzidine (DAB) as chromo- 
gen. Sections were counterstained with hematoxylin for 
15 seconds before checked under microscope. Normal 
liver tissues was used as positive control, while negative 
control was done using the same tissue (normal liver), 
omitting the primary antibody. 

Immunohistochemical analysis 

Only cytoplasmic or cytoplasmic and nuclear reactivity 
was considered as positive staining for arginase-1. 
For HepPar-1; positivity was defined as coarsely granu- 
lar cytoplasmic staining that could not be confused 
with background staining or endogenous peroxidase 
staining. Immunoreactivity was semiquantitatively scored 
by 2 pathologists. The intensity of immunostaining was 
scored as 0 (no staining), 1+ (weak staining), and 2+ 
(strong staining). Furthermore, the pattern of staining 
(diffuse or focal) was recorded. Focal staining was defined 
as reactivity in <10% of tumor or lesional cells [6]. 

Statistical analysis 

Statistical analysis was carried out using Statistical 
Package for Social Science (SPSS 15.0.1 for windows; 
SPSS Inc, Chicago, IL, 2001). Qualitative variables are 
expressed as frequencies and percents. Chi square test 
and Fisher s exact test was used to examine the relation- 
ship between categorical variables. McNemar test was 
used to assess the statistical significance of the difference 
between both immunomarkers for the studied cases. The 
equation used for sensitivity of diagnostic measures was: 
True positive by the test/ (True positive by the test + false 
negative by the test) and for specificity; the equation was 
true negative by the test/ (true negative by the test + false 
positive by the test). Positive predictive value (PPV) is 
calculated as true positive by test/all positive by the test 
(True positive by the test + False Positive by the test). 
Negative predictive value (NPV) is calculated as true 
negative by test/all negative by the test (True negative by 



the test + false negative by the test) with histologic diag- 
nosis designated as the gold standard. 

Results 

Clinicopathologic features 

The fifty cases of HCC were graded as 11 well differ- 
entiated, 30 moderately differentiated, and 9 poorly 
differentiated . All HCC cases are associated with hepa- 
titis C viral (HCV) infection. Forty-two of the 50 HCC 
cases were surgically resected specimens and had adja- 
cent non-neoplastic liver tissues that revealed cirrhotic 
liver tissues and 8 were needle core biopsies. Only two 
cases of HCC were biopsies of metastatic sites (adrenal 
gland and chest wall) and the remaining were primary to 
the liver. The 38 cases of metastatic carcinoma to the 
liver including 25 from colon, 6 from stomach, 1 from 
gall bladder and 2 each from pancreas, kidney and lung. 
The non-neoplastic liver tissues adjacent to metastatic 
carcinomas were detected in 9 cases and revealed no 
pathological abnormalities. 

Immunohistochemical findings 

Immunohistochemical expressions of arginase-1 and 
HepPar-1 in all the studied cases were summarized 
in Tables 1, 2, 3 and 4 in addition to Figures 1, 
2, 3, 4 and 5. 

All normal liver tissues (no=10), non- neoplastic cir- 
rhotic liver tissues adjacent to HCC (no =42) as well as 
those adjacent to MC (no= 9) showed diffuse and strong 
(2+) immunostaining for both arginase-1 and HepPar-1. 

Arginase-1 demonstrated positive immunoreactivity 
in 42 of 50 (84%) cases of HCC compared with 35 of 
50 (70%) for HepPar-1. Positive arginase -1 and HepPar-1 
expression was present in all 11 cases (100%) of well- 
differentiated HCC. However; arginase -1 immunostain- 
ing was positive in 27 of 30 (90%) cases of moderately 
differentiated HCC and 4 of 9 (44.4%) cases of poorly dif- 
ferentiated HCC compared with 22 of (30) (73.3%) and 2 
of (9) (22.2%) for HepPar-1 respectively . In all studied 
HCC cases, there were no cases that were positive for 
HepPar-1 with concurrent negative arginase-1 staining, 
while 7 HCC cases showed arginase-1 staining but were 
negative for HepPar-1. 

Only one of 38 (2.6%) cases of MC and one of 
12 (8.3%) cases of CC showed positive immunoreactivity 
for arginase-1 and the staining was focal and weak. 
In contrast, HepPar-1 immunoreactivity was detected in 
6 of 38 (15.8%) cases of MC and in 2 of 12 (16.7%) 
cases of CC. 

Among all HCC cases, arginase -1 showed a signifi- 
cantly higher sensitivity for diagnosis of HCC (84%) com- 
pared to HepPar -1 (70%) (p=0.016). Within the different 
grades of HCC; the sensitivities of arginase-1 in well, 
moderately, and poorly differentiated HCCs are 100%, 



Radwan and Ahmed Diagnostic Pathology 2012, 7:149 
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Page 4 of 12 



Table 1 Clinicopathological features and the expressions of arginase-1 & HepPar-1 in all studied tumorous cases 
(no=100) 



No 


Aq6 




nibujiuy ii_di uiayinjbib ui iiic lumui 


Arninaco-1 avnrQccinn 
Ml y 1 1 Idbc I caJJi trbblLM 1 


UanPar-1 avnraccinn 
ncJJrdl I CAJJI trbblLM I 


1 


53 


M 


WpII HiffprpntiatprI \-\CC 
vvcii u 1 1 1 ci ci i Lid lcu n^.v_ 


2+ 


2+ 


2 


zm 


M 


WpII Hiffprpnti^tPrl YACC 
Well Ull Iclcl ILIdLcU nV_v^ 


1 _|_ 


1 _|_ 


j 




M 


WpII Hiffprpnti^tPrl YACC 
Well Ull Iclcl ILIdLcU nV_V_ 


2+ 


2+ 


4 


59 


p 


WpII HiffprpntipitpH YACC 

Well U 1 1 1 cl cl 1 Lid LcU nLL 


2+ 


2+ 


5 


60 


M 


WpII HiffprpntipitpH YACC 

Well U 1 1 1 cl cl 1 Lid LcU nLL 


1 _|_ 


2+ 


6 


49 


M 


WpII HiffprpntiatpH \-\CC 

WCII U 1 1 1 Cl Cl 1 Lid LCU n^.^. 


2+ 


1 _|_ 


7 


^3 


p 


WpII Hiffprpnti^tPrl YACC 
Well Ull Iclcl ILIdLcU nV_v^ 


2+ 


1 _|_ 


Q 
o 


jj 


M 


WpII HiffprpntipitpH YACC 
Well Ull Iclcl ILIdLcU nV_V_ 


2+ 


2+ 


9 


48 


M 


WpII Hiffprpnti^tprl YACC 

Well U 1 1 1 cl cl 1 Lid LcU nLL 


2+ 


2+ 


1 0 


50 


p 


WpII HiffprpntiatpH YACC 

Well U 1 1 1 cl cl 1 Lid LcU nLv, 


1 _|_ 


1 _|_ 


1 1 


^9 

jz 


r 


WpII HiffprpntisfpH YACC 
Well Ull Iclcl ILIdLcU n^_^_ 


zt 


zt 


1 2 


zlQ 


M 


K/lnHpratpl\/ HiffprpntiafpH YACC 
IVIUUcldLciy Ull Iclcl ILIdLcU n^.v_ 


n 

u 


n 

u 


1 3 


59 


M 


K/lnHprstpK/ HiffprpntisfpH YACC 
IVIUUcldLciy U 1 1 1 cl Cl 1 Lid LcU nv^_v^_ 


2+ 


2+ 


1 4 


62 


M 


K/lnHprstpK/ HiffprpntisfpH YACC 
IVIUUcldLciy U 1 1 1 cl cl 1 Lid LcU nLL 


1 _|_ 


1 _|_ 


1 5 


59 


M 


K/lnHprptpK/ rliffprpntipitprl Hf~f~ 

IVIUUCIdLCiy U 1 1 1 Cl Cl 1 Lid LCU nLL 


1 _|_ 


1 _|_ 


1 ft 

I D 


^9 

JZ 


p 


K/lnHpratpl\/ HiffprpntiafpH YACC 
IVIUUcldLciy Ull Iclcl ILIdLcU n^.^_ 


2+ 


2+ 


1 7 


JJ 


M 


K/lnHpratpl\/ Hiffprpntiafprl YACC 
IVIUUcldLciy Ull Iclcl ILIdLcU n^.^_ 


1 _|_ 


1 _|_ 


1 8 


56 


p 


K/lnHprstpK/ HiffprpntisfpH YACC 
IVIUUcldLciy U 1 1 1 cl cl 1 Lid LcU nLL 


2+ 


2+ 


1 9 


61 


M 


K/lnHprstpK/ HiffprpntisfpH YACC 
IVIUUcldLciy U 1 1 1 cl cl 1 Lid LcU nLL 


1 _|_ 


Q 


20 


60 


M 


MnHprpfpIv Hiffprpnti^tpH Hf~f~ 

IVIUUCIdLCiy U 1 1 1 Cl Cl 1 Lid LCU nLL 


2+ 


] _)_ 


21 


Dy 


M 


K/lnHpratpl\/ HiffprpntiafpH YACC 
IVIUUcldLciy Ull Iclcl ILIdLcU n^.^_ 


2+ 


] _|_ 


22 


J I 


M 


K/lnHpratpl\/ Hiffprpntiafprl 1— \CC 
IVIUUcldLciy Ull Iclcl ILIdLcU n^.^_ 


n 

u 


n 

u 


23 


55 


p 


K/lnHprstpK/ HiffprpntisfpH YACC 
IVIUUcldLciy U 1 1 1 cl Cl 1 Lid LcU nLL 


2+ 


2+ 


24 


61 


M 


K/lnHprstpK/ HiffprpntisfpH YACC 
IVIUUcldLciy U 1 1 1 cl cl 1 Lid LcU nLL 


1 _|_ 


1 _|_ 


25 


49 


M 


K/lnHprstpK/ HiffprpntisfpH YACC 
IVIUUcldLciy U 1 1 1 cl cl 1 Lid LcU nv_L 


2+ 


2+ 


ZD 


JJ 


M 


K/lnHpratpl\/ HiffprpntiafpH 1— Iftft 
IVIUUcldLciy Ull Iclcl ILIdLcU n^.v_ 


1 _|_ 


n 

u 


27 


^zt 


M 


K/lnHpratpl\/ HiffprpntiafpH 1— Iftft 
IVIUUcldLciy Ull Iclcl ILIdLcU n^.v_ 


2+ 


2+ 


28 


59 


p 


K/lnHprstpK/ HiffprpntisfpH l-lftft 
IVIUUcldLciy U 1 1 1 cl cl 1 Lid LcU nLL 


2+ 


2+ 


29 


62 


M 


K/lnHprstpK/ HiffprpntisfpH l-lftft 
IVIUUcldLciy U 1 1 1 cl cl 1 Lid LcU nLL 


2+ 


2+ 




fti 

O I 


f VI 


K/lnrlpratpK/ Hiffprpntiatprl YACC 
IVIUUcldLciy Ull Iclcl ILIdLcU n^_^_ 


1 T 


n 
u 




JU 


M 


K/lnHpratpl\/ HiffprpntiafpH 1— \CC 
IVIUUcldLciy Ull Iclcl ILIdLcU n^.v_ 


] _|_ 


] _|_ 


DZ 


ZLR 

to 


M 


K/lnHprafpK/ HiffprpntiafpH 1— Iftft 
IVIUUcldLciy Ull Iclcl ILIdLcU n^.^_ 


2+ 


] _|_ 


33 


52 


p 


K/lnHprstpK/ HiffprpntisfpH l-lftft 
IVIUUcldLciy U 1 1 1 cl cl 1 Lid LcU nv^_v_ 


2+ 


2+ 


34 


60 


p 


MnHprpfpIv Hiffprpnti^tprl Hf~f~ 

IVIUUCIdLCiy U 1 1 1 Cl Cl 1 Lid LCU nv_v_ 


1 + 


o 


3^ 


^9 

JZ 


M 


K/lnHpratpl\/ Hiffprpntiafprl 1— Iftft 
IVIUUcldLciy Ull Iclcl ILIdLcU n^.^_ 


2-j- 


2+ 


3ft 

DO 


^ft 

JO 


M 


K/lnHpratpl\/ HiffprpntiafpH 1— \CC 
IVIUUcldLciy Ull Iclcl ILIdLcU n^.^_ 


1 _|_ 


1 _|_ 


37 


57 


M 


K/lnHprstpK/ HiffprpntisfpH YACC 
IVIUUcldLciy U 1 1 1 cl cl 1 Lid LcU nLL 


Q 


Q 


38 


58 


p 


K/lnrlprpitpl\/ rliffprpntipitprl YACC 
i viuuci a iciy ui 1 1 ci ci i ua lcu i 


2+ 


2+ 


39 


65 


M 


Moderately differentiated HCC 


2+ 


2+ 


40 


55 


M 


Moderately differentiated HCC (metastatic to chest wall) 


1 + 


1 + 


41 


48 


M 


Moderately differentiated HCC (metastatic to adrenal gland) 


1 + 


0 


42 


53 


M 


Poorly differentiated HCC 


0 


0 


43 


57 


F 


Poorly differentiated HCC 


1 + 


0 


44 


62 


M 


Poorly differentiated HCC 


0 


0 


45 


65 


M 


Poorly differentiated HCC 


1 + 


1 + 


46 


50 


F 


Poorly differentiated HCC 


1 + 


0 



Radwan and Ahmed Diagnostic Pathology 2012, 7:149 
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Page 5 of 12 



Table 1 Clinicopathological features and the expressions of arginase-1 & HepPar-1 in all studied tumorous cases 
(no=100) (Continued) 



47 


54 


M 


Poorly differentiated HCC 


0 


0 


48 


56 


M 


Poorly differentiated HCC 


0 


0 


49 


52 


M 


Poorly differentiated HCC 


1 + 


1 + 


50 


60 


M 


Poorly differentiated HCC 


0 


0 


51 


62 


F 


Metastatic colonic carcinoma 


0 


0 


52 


54 


M 


Metastatic colonic carcinoma 


0 


0 


53 


50 


F 


Metastatic colonic carcinoma 


0 


0 


54 


45 


F 


Metastatic colonic carcinoma 


0 


0 


55 


48 


M 


Metastatic colonic carcinoma 


0 


0 


56 


50 


F 


Metastatic colonic carcinoma 


0 


1 + 


57 


57 


M 


Metastatic colonic carcinoma 


0 


0 


58 


49 


F 


Metastatic colonic carcinoma 


0 


0 


59 


55 


F 


Metastatic colonic carcinoma 


0 


2+ 


60 


50 


M 


Metastatic colonic carcinoma 


0 


0 


61 


52 


M 


Metastatic colonic carcinoma 


0 


0 


62 


60 


M 


Metastatic colonic carcinoma 


0 


0 


63 


51 


M 


Metastatic colonic carcinoma 


0 


0 


64 


55 


F 


Metastatic colonic carcinoma 


0 


2+ 


65 


50 


M 


Metastatic colonic carcinoma 


0 


0 


66 


49 


F 


Metastatic colonic carcinoma 


0 


0 


67 


55 


F 


Metastatic colonic carcinoma 


0 


0 


68 


51 


F 


Metastatic colonic carcinoma 


0 


0 


69 


53 


M 


Metastatic colonic carcinoma 


0 


0 


70 


55 


M 


Metastatic colonic carcinoma 


0 


0 


71 


50 


M 


Metastatic colonic carcinoma 


0 


0 


72 


52 


M 


Metastatic colonic carcinoma 


0 


0 


73 


56 


F 


Metastatic colonic carcinoma 


0 


0 


74 


46 


M 


Metastatic colonic carcinoma 


0 


0 


75 


50 


M 


Metastatic colonic carcinoma 


0 


0 


76 


52 


M 


Metastatic gastric carcinoma 


0 


2+ 


77 


49 


F 


Metastatic gastric carcinoma 


0 


0 


78 


55 


M 


Metastatic gastric carcinoma 


0 


2+ 


79 


56 


M 


Metastatic gastric carcinoma 


0 


0 


80 


50 


M 


Metastatic gastric carcinoma 


0 


1 + 


81 


52 


F 


Metastatic gastric carcinoma 


0 


0 


82 


53 


M 


Metastatic renal cell carcinoma 


0 


0 


83 


55 


F 


Metastatic renal cell carcinoma 


0 


0 


84 


61 


M 


Metastatic pancreatic carcinoma 


1 + 


0 


85 


62 


M 


Metastatic pancreatic carcinoma 


0 


0 


86 


60 


M 


Metastatic lung carcinoma 


0 


0 


87 


62 


F 


Metastatic lung carcinoma 


0 


0 


88 


55 


M 


Metastatic gall bladder carcinoma 


0 


0 


89 


60 


M 


Cholagiocarcinoma 


1 + 


0 


90 


64 


M 


Cholagiocarcinoma 


0 


1 + 


91 


59 


M 


Cholagiocarcinoma 


0 


2+ 


92 


64 


F 


Cholagiocarcinoma 


0 


0 


93 


62 


F 


Cholagiocarcinoma 


0 


0 



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Table 1 Clinicopathological features and the expressions of arginase-1 & HepPar-1 in all studied tumorous cases 
(no=100) (Continued) 



94 


58 


M 


Cholagiocarcinoma 


0 


0 


95 


62 


F 


Cholagiocarcinoma 


0 


0 


96 


60 


M 


Cholagiocarcinoma 


0 


0 


97 


59 


F 


Cholagiocarcinoma 


0 


0 


98 


65 


M 


Cholagiocarcinoma 


0 


0 


99 


62 


M 


Cholagiocarcinoma 


0 


0 


100 


60 


M 


Cholagiocarcinoma 


0 


0 



M= male, F= female, HCC= hepatocellular carcinoma, HepPar-1=hepatocyte paraffin antigen-1,(0) =no staining, (1+) =weak staining, (2+) =strong staining. 



90%, and 44.4%, respectively, whereas, in comparison, 
HepPar- 1 demonstrated sensitivities of 100%, 73.3%, and 
22.2% for well, moderately, and poorly differentiated 
tumors, respectively. There was no significant difference 
between arginase -1 and HepPar- 1 as regards their sensi- 
tivities in diagnosis of well or poorly differentiated HCC, 
while for moderately differentiated HCC cases; arginase 
-1 showed a significantly higher sensitivity than HepPar- 1 
(p=0.001). 

The specificity of arginase-1 for diagnosis of HCC was 
higher (96%) than that of HepPr -1 (84%); nevertheless, 
this was not statistically significant (p= 0.109). The posi- 
tive predictive value (PPV) of arginase-1 for distinguish- 
ing HCC from MC and CC was higher (95.5%) than that 
observed with HepPar- 1(81.4%). Also, the negative pre- 
dictive value (NPV) for arginase-1 (85.7%) in distin- 
guishing HCC from MC and CC was better than that of 



HepPar- 1(73.7%). Howerver, the combination of both 
immunomarkers for the diagnosis of HCC, raised the 
specificity to 100% as shown in Table 4. 

Discussion 

The most commonly encountered differential diagnostic 
challenge in the liver is HCC versus intrahepatic cholan- 
giocarcinoma or metastatic adenocarcinoma [7]. Some 
of these diagnostic challenges can be attributed to: a) 
The liver represent one of the three most common sites 
of metastasis, b) HCCs may show a variety of histologic 
patterns, mimicking a wide variety of malignant tumors. 
In addition, a number of metastatic tumours, notably from 
the breast, pancreas, kidney and adrenals may mimic the 
trabecular, liver-like pattern of HCC, c) Cholangiocarci- 
noma and HCC often share overlapping morphologic 
appearances, d) Complicating the diagnostic process is 



Table 2 Summary of immunohistochemical expression of arginase-1 and HepPar- 1 in all the studied cases 



Arginase-1 HepPar-1 





Negative 


Positive 


Total 
positive 


Negative 


Positive 


Total 
positive 




0 (%) 


1+(%) 


2+(%) 


No (%) 


0(%) 


1+(%) 


2+(%) 


No (%) 


Hepatocellular carcinoma (HCC), (no=50): 


8(16) 


19 (38) 


23 (46) 


42 (84) 


15 (30) 


16 (32) 


19 (38) 


35 (70) 


well differentiated (no=11) 


0(0) 


3 (27.3) 


8 (72.7) 


11 (100) 


0(0) 


4 (36.4) 


7 (63.6) 


11 (100) 


moderately differentiated (no=30) 


3 (10) 


12 (40) 


15 (50) 


27 (90) 


8 (26.7) 


10 (33.3) 


12 (40) 


22 (73.3) 


poorly differentiated (no=9) 


5 (55.6) 


4 (44.4) 


0(0) 


4 (44.4) 


7 (77.8) 


2 (22.2) 


0(0) 


2 (22.2) 


Metastatic carcinoma (MC) (no=38) 


37 (97.4) 


1 (2.6) 


0(0) 


1 (2.6) 


32 (84.2) 


2 (5.3) 


4 (10.5) 


6 (15.8) 


Colonic (no=25) 


25 (100) 


(0) 


0(0) 


0(0) 


22 (88) 


1 (4) 


2(8) 


3(12) 


Gastric (no=6) 


6 (100) 


0(0) 


0(0) 


0(0) 


3(50) 


1 (16.7) 


2 (33.3) 


3(50) 


Renal cell carcinoma (no=2) 


2 (100) 


0(0) 


0(0) 


0(0) 


2 (100) 


0(0) 


0(0) 


0(0) 


Pancreas (no=2) 


1 (50) 


1 (50) 


0(0) 


1 (50) 


2 (100) 


0(0) 


0(0) 


0(0) 


Lung (no=2) 


2 (100) 


0(0) 


0(0) 


0(0) 


2 (100) 


0(0) 


0(0) 


0(0) 


Gall bladder (no=1) 


1 (100) 


0(0) 


0(0) 


0(0) 


1 (100) 


0(0) 


0(0) 


0(0) 


Cholangiocarcinoma (no=12) 


11 (91.7) 


1 (8.3) 


0(0) 


1 (8.3) 


10 (83.3) 


1 (8.3) 


1 (8.3) 


2(16.7) 


Non-neoplastic cirrhotic liver tissues 
adjacent to HCC(no=42) 


0(0) 


0(0) 


42 (100) 


42 (100) 


0(0) 


0(0) 


42 (100) 


42 (100) 


Non-neoplastic liver tissue adjacent to MC (no= 9) 


0(0) 


0(0) 


9 (100) 


9 (100) 


0(0) 


0(0) 


9 (100) 


9 (100) 


Normal liver tissues (no=10) 


0(0) 


0(0) 


10 (100) 


10 (100) 


0(0) 


0(0) 


10 (100) 


10 (100) 



HepPar-1=hepatocyte paraffin antigen-1. 



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Table 3 Immunohistochemical expression of arginase-1 and HepPar-1 according to the pattern of staining in all 
positive cases 







Arginase-1 






HepPar-1 






Focal No (%) 


Diffuse 

No (0/ n \ 


Total 

fJUblUVc 


Focal 

No (0/ n \ 


Diffuse 

No (0/ n \ 


Total 

rt/tci+ii/a 
[JUblU Vc 


ncjJd luutriiuidi Cdiuiiiuiiid V nv.v»j , ^nu — juj. 


1 0 (23 8) 


32 (76 2) 


42 


1 5 (42 9) 


20 (57 1) 


35 


\a/oII H ifforonti atoH (nn — 1 1^ 
Well U I II Cl cl I Lid LcU \\ IU — I I ) 


n (0) 
u yu) 


1 1 (1 00) 


1 1 


i \y.\) 


l U \y\j.y) 


1 1 


lllUUcldLcly Ull Iclcl ILIdLcU \\W — jUJ 


9. (1Q £\ 
O \Zy.\D) 


1 Q (70 A\ 


27 


1 n T\ 

I U \DD.D) 


1 1 (AO) 
1 Z \f-t\J) 


22 


pUUIly Ull Icltrl 1 Lid LcU \\ IU— y) 


1 (W\ 

z puj 


z pu; 


4 


1 (11 1\ 
Z \ZZ.Z) 


n ith 

u [yi) 


2 


Metastatic carcinoma (MC) (no=38): 


1 (100) 


0(0) 


1 


1 (16.7) 


5 (83.3) 


6 


Colonic (no=25) 


(0) 


0(0) 


0 


1 (33.3) 


2 (66.7) 


3 


Gastric (no=6) 


0(0) 


0(0) 


0 


0(0) 


3 (100) 


3 


Renal cell carcinoma (no=2) 


0(0) 


0(0) 


0 


0(0) 


0(0) 


0 


Pancreas (no=2) 


1 (100) 


0(0) 


1 


0(0) 


0(0) 


0 


Lung (no=2) 


0(0) 


0(0) 


0 


0(0) 


0(0) 


0 


Gall bladder (no=1) 


0(0) 


0(0) 


0 


0(0) 


0(0) 


0 


Cholangiocarcinoma (no=12) 


1 (100) 


0(0) 


1 


1 (50) 


1 (50) 


2 


Non-neoplastic cirrhotic liver tissues 
adjacent to HCC(no=42) 


0(0) 


42 (100) 


42 


0(0) 


42 (100) 


42 


Non-neoplastic liver tissue adjacent to 
MC (no= 9) 


0(0) 


9 (100) 


9 


0(0) 


9 (100) 


9 


Normal liver tissues (no=10) 


0(0) 


10 (100) 


10 


0(0) 


10 (100) 


10 



HepPar-1=hepatocyte paraffin antigen-1. 



that pathologists are frequently asked to handle and 
diagnose tiny liver needle core biopsies with various 
biopsy artifacts [9,22]. A limited number of diagnostically 
useful immunohistochemical markers have been applied 
in an attempt to differentiate HCC from liver metastases 
or cholangiocarcinoma including; HepPar-1, polyclonal 
carcinoembryonic antigen (CEA), and CD 10, with alfa- 
fetoprotein (AFP) and glypican-3 labeling some HCCs [6]. 
However, the utility of each of these markers has signifi- 
cant diagnostic limitations [7] . 

A recent study of Hajosi-Kalcakosz et al. [23] pub- 
lished in 2012 investigated enhancer of zeste homologue 
2 (EZH2) as a new marker of HCC. They reported that 
EZH2 was detected by immunohistochemistry in nearly 
all the investigated HCC, CC, hepatoblastoma, meta- 
static liver tumors and several other childhood cancers. 
On the contrary, none of the hepatocellular or biliary 
adenomas, high grade dysplastic or cirrhotic nodules 
was positive. Thus, this study concluded that EZH2 is a 
sensitive and reliable immune marker of hepatocellular 
carcinoma, compared to non-malignant hepatocellular 
lesions. However, EZH2 is not specific for HCC, since 
almost all the investigated malignant liver tumors were 
positive as well regardless of their histogenesis. Conse- 
quently, this marker does not provide help in differenti- 
ating the specific histogenesis of liver tumors, but it may 
well be very useful to differentiate malignant hepatocel- 
lular and cholangiocellular tumors from benign tumors 
and reactive lesions. 



Moreover, special stains, such as reticulin stain and 
CD34 immunostain, are very helpful in the diagnosis of 
well differentiated HCC. Most studies have shown that 
absent or decreased reticulin stain or an abnormal re- 
ticulin pattern with widened trabeculae is reliable for the 
diagnosis of well-differentiated HCC. However, Hong 
et al. [24] reported two cases of well- differentiated HCC 
with an unusual reticulin staining pattern in their pri- 
mary biopsies. They suggested that HCC may have di- 
verse reticulin patterns in different portions of the tumor. 
In a small specimen, such as core biopsy, if only the por- 
tion of tumor with well preserved reticulin network is 
present, the diagnosis can be challenging. Thus, it is 
important to recognize the presence of different reticulin 
staining patterns in the evaluation of small biopsies for 
the diagnosis of HCC. 

Arginase-1 has been described in recent literature as a 
new potential immunohistochemical marker of hepato- 
cellular differentiation [6]. Only few studies investigated 
arginase -1 expression in HCC and most of these 
reports performed on fine needle aspiration cytology 
[5,25,26] with some variation in their interpretations as 
regards its sensitivity and specificity. Therefore; the pri- 
mary purpose of the current study was to examine the 
immunohistochemical staining of arginase-1 in cases of 
HCC, metastatic carcinoma involving the liver and cho- 
langiocarcinoma as compared to HepPar-1. This is in an 
attempt to further define its diagnostic utility as a reli- 
able positive marker in differentiating these tumors. 



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Table 4 Sensitivity, specificity, positive and negative 
predictive value of arginase-1, HepPar-1 for HCC 
diagnosis 





Sensitivity 


Specificity 


PPV 


NPV 


Arginase-1 


84% 


96% 


95.5% 


85.7% 


HepPar-1 


70% 


84% 


81.4% 


73.7% 


Arginase-1 or HepPar-1 


84% 


80% 


88.8 


83.3% 


Arginase-1 and HepPar-1 


70% 


100% 


100% 


76.9% 



HCC=hepatocellular carcinoma; MC=metastatic carcinoma; 
CC=cholangiocarcinoma; PPV=positive predictive value; NPV=negative 
predictive value; HepPar-1=hepatocyte paraffin antigen-1. 



HepPar- 1 was selected to be compared with this new 
marker as it is conventially used and has been found to 
be overrated as a hepatoma marker. The present study 
examined arginase-1 and HepPar- 1 expression in 50 HCC 
cases, 38 cases of metastatic carcinomas to the liver from 
varying sites, 12 cases of cholangiocarcinoma and 10 spe- 
cimens of normal liver tissues. In addition, the non- 
neoplastic liver tissues adjacent to HCC or metastatic 
carcinomas were also investigated. 

The results revealed that arginase -1 showed a signifi- 
cantly higher overall sensitivity for diagnosis of HCC 
(84%) compared to HepPar -1 (70%). This confirm the 
conclusion of the previous studies [5,6,25-27]. It is worth 
mentioning that there were no cases were positive for 
HepPar- 1, with concurrent negative arginase-1 staining. 
In addition, arginase-1 showed more diffuse staining in 
HCC (76.2%) than HepPar-1 (57.1%). This makes inter- 
pretation of arginase -1 easier especially in limited liver 
biopsies. 

Furthermore, arginase-1 gave a sensitivity of 100%, 
90%, and 44.4% in well, moderately, and poorly differen- 
tiated HCCs, respectively, whereas, in comparison, 
HepPar- 1 demonstrated sensitivities of 100%, 73.3%, and 
22.2% for well, moderately, and poorly differentiated 
tumors, respectively. Therefore, arginase-1 showed bet- 
ter sensitivity compared with HepPar- 1 in identifying 



higher grade HCC. This is relatively in accordance with 
the original paper describing the antibody of Yan et al. 
[6] who found more marked difference between both 
immunomakers in poorly differentiated HCCs, in which 
the sensitivities of arginase-1 and HepPar- 1 were 85.7% 
and 46.4%, respectively. This finding is very useful because 
one of the most frequent diagnostic challenges facing a 
pathologist examining liver focal lesion is distinguishing 
between poorly differentiated HCC from a metastasis, 
especially in small biopsy specimen. The lower diagnostic 
sensitivity in our study as compared to that of Yan et al. 
[6] may be because of the smaller sample size. In contrast, 
Timek et al. [25] failed to demonstrate a better sensitivity 
of arginase-1 for higher-grade HCC compared with 
HepPar- 1 and they explained that by the small sampling 
of the cytologic specimens in the moderately to poorly 
differentiated HCC category (n =7), limited amount of 
sample for each case, and patchy/focal staining for 
arginase-1 in higher-grade HCC. 

Moreover, we observed diffuse and strong immunos- 
taining for both arginase-1 and HepPar- 1 in the non- 
neoplastic cirrhotic liver tissues adjacent to HCC as 
well as those adjacent to MC. This supports the study of 
Fujiwara et al. [5] and Timek et al. [25] who reported that 
arginase-1 has no role in distinguishing well-differentiated 
hepatocellular carcinoma from benign hepatic lesions. 

Two very recent studies examined the immunohisto- 
chemical expression of LI cell adhesion molecule 
(L1CAM) [28] and SOX9 [29] in HCC cases and their 
adjacent non- neoplastic liver tissues and they reported 
that immunoreactivity of these markers was significantly 
increased in substantial proportion of HCC cases com- 
pared with their adjacent non- neoplastic liver tissue. 
Additionally, they suggested that L1CAM expression in 
HCC was significantly correlated with the advanced 
tumor progression and was an independent poor prog- 
nostic factor for both overall survival and disease-free 
survival in patients with HCC. Furthermore, SOX9 over- 
expression in HCC tissues is of predictive value on tumor 




Figure 1 A case of moderately differentiated hepatocellular carcinoma (A, H&E,original magnification x400) with strong and diffuse 
arginase-1 staining (B; immuoperoxidase, original magnification x400) and focal HepPar- 1 immunostaining (C; immuoperoxidase, 
original magnification x400). 



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Page 9 of 12 




progression and poor prognosis. Moreover, Schmilovitz- 
Weiss et al [30] reported that squamous cellular carcin- 
oma antigen (SCCA) is overexpressed in HCC and it is 
associated with tumor differentiation, cell proliferation 
and apoptosis. The results of their study confirm a po- 
tential association of negative SCCA expression with 
other markers of poor outcome in HCC. 

In our study, the specificity of arginase-1 for diagnosis 
of HCC was higher (96%) than that of HepPar -1 (84%). 
Only one case of pancreatic adenocarcinoma out of 
38 (2.6%) cases of MC and one of 12(8.3%) cases of 
CC showed positive immunoreactivity for arginase-1. 
However, the staining was focal and weak in these two 
positive cases. In contrast, HepPar- 1 immunoreactivity 
was detected in 6 of 38 (15.8%) cases of MC (3 from 
colon and 3 from stomach) and in 2 of 12 (16.7%) cases 
of CC. Although, neither arginase-1 nor HepPar- 1 immu- 
nostaining demonstrated 100% diagnostic specificity to 
distinguish HCC from MC in the liver and CC, our 
analysis of the combination of both immunomarkers 
among all studied tumors, raised the diagnostic specifi- 
city for HCC to 100% if both showed positive immu- 
nostainings. This high specificity of arginase-1 and 
HepPar- 1 combination because the staining patterns of 



both immunomarkers in adenocarcinomas were mutu- 
ally exclusive (i.e. arginase-1 - positive adenocarcinomas 
always lacked HepPar- 1 immunoreactivity and vice 
versa) [5]. 

These findings are in agreement with the study of 
Fujiwara et al. [5] which showed that arginase-1 is not 
entirely specific for hepatic differentiation, as immunor- 
eactivity can be identified in adenocarcinomas, particu- 
larly of pancreatic origin. The authors reported that it is 
not surprising to find a subset of the pancreatic adeno- 
carcinomas included in their analysis demonstrated 
arginase-1 immunoreactivity. This is because a recent 
analysis of arginase-1 immunohistochemical expression 
in rats demonstrated that it was expressed at high levels 
in the liver and at moderate levels in the pancreas [18]. 
Moreover, Yan et al. [6] found that only one case of 
prostatic adenocarcinoma demonstrated arginase-1 immu- 
noreactivity. Of note, their study did not include pan- 
creatic adenocarcinomas in their analysis. In contast, 
Timek et al. [25] and McKnight et al. [26] reported 
negativity of arginase-1 in all their cases of MC. 

The positive immunostaining of HepPar- 1 in our 6 cases 
of MC (3 from colon and 3 from stomach) was in con- 
cordance with the results of Yan et al. [6] who detected 




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Figure 4 A case of metastatic colonic adenocarcinoma to the liver (A,D; H&E,original magnification x200,x400) showed negative 
arginase-1 immunostaining (B,E; immuoperoxidase, original magnification x200, x400), and stong and diffuse staining with HepPar-1 
(C,F; immuoperoxidase, original magnification x200, x400). 

) 



HepPar-1 reactivity in 2 colonic adenomas, 8 colonic 
adenocarcinomas, 2 pulmonary adenocarcinomas, 1 chro- 
mophobe RCC, and 9 gastric adenocarcinomas (47.4% 
of cases). HepPar-1 immunoreactivity in gastric adeno- 
carcinomas is reported in previous studies in which it was 
expressed in 47% to 83% of gastric cancers [10,13,31]. 
Moreover, Timek et al. [25] reported that the expression 
of HepPar-1 in nonhepatocellular tumors is well docu- 
mented in the literature and they assumed that caution 
should be taken when using HepPar-1 to confirm a diag- 
nosis of HCC. 

In our study, out of 12 cases of CC, only one (8.3%) 
was positive for arginase-1, while 2 (16.7%) were positive 
for HepPar-1. This supports the study of Yan et al. as 



regards arginase -1 reactivity [6]. In addition, Fujiwara 
et al. [5] reported negative immunoreactivity in all their 
cases for both immunomarkers. However, the positivity 
of HepPar-1 in our study is consistent with previous 
studies [14,22,32]. Shiran et al. [22] claimed that the 
presence of this occasional positivity should not be sur- 
prising considering the common progenitor cell of HCC 
and CC [14]. On the contrary, Iida et al. [33] concluded 
that HepPar-1 was rarely but definitely expressed in hilar 
and peripheral intrahepatic CC, while arginase-1 was 
expressed at a high rate in both hilar and peripheral 
intrahepatic CC, irrespective of their histology. They 
assumed that care should be taken when using arginase-1 
as a hepatocyte marker for distinguishing between a 




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poorly differentiated hepatocellular carcinoma and a 
mass-forming peripheral intrahepatic CC showing the 
histology of poorly differentiated adenocarcinoma. 

One of the important findings in the present study 
was that arginase-1 showed diffuse and strong nuclear 
reactivity along with cytoplasmic staining which was 
observed more in some HCC cases and their adjacent 
non-neoplastic cirrhotic liver tissues compared with 
other studied cases. It could be explained as all our 
HCC cases are associated with HCV. This possible 
explanation is supported by the findings of Cao et al. [34] 
who reported that elevated arginase-1 staining is asso- 
ciated with chronic HCV infection as they found that 
arginase-1 expression was elevated in more than 75% 
of HCV infected liver samples compared to paired 
HCC from the same patients (> 33% positive) and to 
uninfected liver tissues (0% positive). The authors sug- 
gested that up-regulated expression of arginase- 1 was 
associated with HCV infected liver, and to a lesser extent 
in tumor, but not in uninfected liver. They assumed that 
an important part of the mechanism whereby HCV regu- 
lates hepatocellular growth and survival may be through 
altering arginine metabolism. However, further studies in 
large scale are worth-while to confirm these observations. 

Conclusions 

In conclusion, the present study demonstrates that 
arginase-1 immunostaining has a higher sensitivity and 
specificity than HepPar-1 for HCC diagnosis. Although 
none of them gives 100% specificity for HCC, but the 
combined use of arginase-1 and HepPar-1 can provide a 
potentially promising tool to improve the accuracy in 
distinguishing HCC from MC and CC. Therefore, from 
the findings of the current and previous few studies 
about arginase-1 immunostaining in HCC, we can expect 
that it will be used as a hepatoma marker in routine sur- 
gical pathology practice. However, further prospective 
studies are recommended to confirm these results. 

Abbreviations 

HCC: Hepatocellular carcinoma; MC: Metastatic carcinoma; 
CC: Cholangiocarcinoma; HCV: Hepatitis C viral; HepPar-1: Hepatocyte 
paraffin antigen-1; H&E: Hematoxylin and eosin; PPV: Positive predictive 
value; NPV: Negative predictive value. 

Competing interests 

The authors declare that they have no competing interests. 
Authors' contributions 

NAR conceived, designed and coordinated the study, evaluated 
immunohistochemistry, performed the statistical analysis, carried out 
photographing and drafted the manuscript. NSA reviewed the histological 
diagnosis, evaluated immunohistochemistry, participated in the study design 
and helped to draft the manuscript. All authors read and approved the final 
manuscript. 

Acknowledgement 

The authors gratefully thank Professor Dr. Thanaa El A Helal, Pathology 
department, Faculty of Medicine, Ain- Shams University for her generous 



support, valuable guidance and fruitful advices throughout the work in this 
study. 

Received: 16 September 2012 Accepted: 17 October 2012 
Published: 30 October 2012 

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doi:1 0.1 1 86/1 746-1 596-7-1 49 

Cite this article as: Radwan and Ahmed: The diagnostic value of 
arginase-1 immunostaining in differentiating hepatocellular carcinoma 
from metastatic carcinoma and cholangiocarcinoma as compared to 
HepPar-1. Diagnostic Pathology 2012 7:149. 



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